Project Summary Immunosenescence of primary lymphatic organs is characterized by a decrease in the self-renewing potential of precursor cells as well as generation of lineage-committed cells affected by age-related changes. The consequences of immune decline are increased susceptibility to infections and cancer, and reduced responses to vaccinations. We recently reported, in collaboration with P1 and P3 of this Program Project (PP), that a hallmark of aging CD34+ bone marrow precursor cells as well as bona fide conventional dendritic cells (cDC) is the presence of an extensively glycated, carbonylated and lipoxidated proteome, which severely compromises their ability to mount effective immune responses. In this new PP period we propose to use a series of biochemical and biophysical approaches to: 1) functionally characterize the effects of different stressors on adaptive immune responses 2) analyze oxidative modifications of the cDC and M? immuno-proteome generated by these stressors 3) analyze the autophagic pathways involved in disposing the oxidatively modified proteome and 4) implement therapeutical intervention aimed at decreasing cellular stressors and restore adaptive immune functions. The overall working hypothesis of this project is that stressor-induced specific protein posttranslational modifications (carbonylation, glycation, lipoxidation) differently affect the cellular immuno-proteome, its autophagic clearance and ultimately the cDC, macrophages (M?) and CD4+ T cell immune functions. In specific Aim 1 we will analyze the effects of cellular stressor on cDC and M? immunoproteome by using qualitative and quantitative mass spectrometry and free flow fractionation analysis of protein aggregates. The autophagic mechanisms use to dispose of the glycated, carbonylated and lipoxidated proteome will be evaluated using CD11c tissue specific null cells for the three autophagic pathways. In specific Aim 2 we will analyze how different stressors, known to induce proteotoxicity and lipotoxicity compromise cDC-mediated adaptive immune responses. In specific Aim 3 we will evaluate a series of genetic, pharmacologic and dietary therapeutic interventions shared with all projects to determine the best approach to decrease proteotoxicity and lipotoxicity on immune cells and restore adaptive immune functions Relevance: This application delves into previously unexplored areas of immunosenescence and immune cells response to stressors commonly encountered in diabetes, metabolic syndrome, prolonged infections and immune cell exhaustion. The overall fundamental goal is to understand the biochemical implication of each stressor on cDC and M? ability to mount adaptive immune responses and to devise therapeutic interventions aimed at restoring cellular functionality and improve immunosenescence.